Amt of restriction enzyme - (Nov/02/2007 )

Hello All,

I just noticed an advertisement by fermentas quick-digest enzymes. They claim 1µl of enzyme mix cuts 1µg of DNA.

I was wondering if such rough equation applies to routinely used restriction enzymes as well. I usually use 20units of Fermentas restriction enzymes for cutting about 3µg of DNA. I would like to know what others use.

Thanks in advance.

-Jiang M-

It would be dependent on the enzyme but most times, the company will tell you that 1U will digest 1 µg DNA in 50 µl volume. I standardly use this ratio when digested DNA and it works.

AussieUSA.

-AussieUSA-

I normally add the enzyme such that in total it should not exceed 10% of the total reaction volume.It always works for me.

-newarray-

Depending on how much DNA you got and how concentrated your enzyme is, just adding an amount in order not to exceed 10% of the reation volume seems like a waste of enzyme. Mostly they come at 5-20 Units/µl which should be enough to cut 5-20 µg of DNA...